Help: How to isolate DNA-Protein crosslinks? - (Sep/11/2006 )

I am working on protein-DNA crosslinks induced by formaldehyde.

I want to know which proteins are crosslinked with DNA.But I have no clear idea about how to isolate DNA-Protein crosslinks from cell cultures. I guess I can get them by collecting DNA. Becasue the crosslinked DNA will be also isolated. Then, I can digest the DNA from DNA-Protein crosslinks, leaving the proteins there. The next step, I can run SDS-PAGE and mass spec to identify the proteins.

Or there is another method called K/SDS. It can precipitate proteins instead of DNA. But it will result in the precipitation of uncrosslinked proteins. That is not what I want.

Does anyone has some idea or protocols about this? Let me make clear my question:How to isolate DNA-Protein crosslinks? Thank you so much.

Kun

-kunkun-

Do you know what proteins do you expect that will bind to the target DNA sequences/and be cross-linked to DNA? If you do, you can immuno-precipitate the protein-DNA complexes.

-genehunter-1-

Thanks a lot.

I expect histone will be crosslinked to DNA induced by chemicals. I need prove this by mass spectrometry. We have no idea about the target DNA sequence. And actually, we donot care about DNA sequence target. We just want to know which proteins will be crosslinked. So I need a general method to isolate DNA-protein crosslinks.

QUOTE (genehunter-1 @ Sep 11 2006, 11:45 AM)

Do you know what proteins do you expect that will bind to the target DNA sequences/and be cross-linked to DNA? If you do, you can immuno-precipitate the protein-DNA complexes.